We have found that a potent tumor promoter, 12-0-tetradecanoyl-phorbol-13-acetate (TPA) can either enhance or inhibit the proliferative response of activated lymphocytes in vitro. The enhancement is due to the ability of TPA to replace the macrophage lymphokine, interleukin 1. Both TPA and the mitogenic lectin Con A are required for DNA synthesis. We found that Con A provides an early stimulation signal and TPA a later one. Con A, but not TPA, can be substituted for by a calcium ionophore. By using TPA, ionophore or lectin in macrophage-free lymphocyte cultures stimulated with the mitogen, Con A, we propose to follow the mechanism of action of stimulation with TPA. The inhibitory effects of TPA occur after a 24-hour exposure and may be due to the differentiation of suppressor cells. We have modified techniques of rosetting with sheep red blood cells to identify three major subclasses of T cells. TPA treatment changes the distribution of cells into these subclasses. The subclasses will be isolated and tested for their effects on lectin-stimulated cells or in mixed lymphocyte cultures. We will determine if any functionally inhibitory subpopulations produce an inhibitory material and attempt to isolate it. Other promoters will be tested in the lymphocyte system to determine if there is a correlation between in vivo promotion and lymphocyte inhibition in vitro. Preliminary experiments indicate that lymphocytes from TPA-treated mice give a reduced response to lectins.